Much evidence suggests that the regulation of the activity of adenosine deaminase is critical for normal cell function and differentiation. The general aim of the proposed research is two-fold: (1) to clarify the subunit structure and interactions which occur in adenosine deaminase; and (2) to understand on a molecular level how the regulation of the enzyme occurs. The basic hypothesis is that regulation is mediated by factors which cause a shift in quaternary structure between a high and a low molecular weight form of the enzyme. Both forms of the enzyme will be highly purified from human and animal sources; their physico-chemical and catalytic properties, including the nature of residues at or near the active site, will be characterized. The low molecular weight species will be isolated from red cells and lymphocytes, and the high molecular weight form from various normal tissues including intestinal epithelial cells at various stages of differentiation. The species of enzyme found in various types of malignant cells will also be isolated and similarly characterized. The purification procedures will involve classical biochemical techniques as well as affinity columns and immunosorbents for rapid isolation of the enzyme from small quantities of tissue. The conversion factor protein which has been reported to promote the low to high molecular weight transition will also be purified from tissues. The relationship of this protein to the three dimensional structure of the high molecular weight enzyme will be elucidated by a variety of physical studies including covalent cross-linking, subunit dissociation, etc. A detailed study of its properties as well as a search for other proteins whose molecular weight it might affect should lead to a clarification of its function. Skin fibroblasts will be cultured from normal subjects and severe combined immunodeficiency disease (SCID) patients. The catalytic and physical properties of the enzyme producing the low residual adenosine deaminase activity in the SCID patient will be compared to those of the enzyme from the normal subjects. These data should clarify the nature of any structural alteration which might account for the reduced activity.